Donnelly CW, Baigent GJ (1986) Method for flow cytometric detection of Listeria monocytogenes in milk. Appl Environ Microbiol 52 :689-695
This report describes a method for the detection of Listeria monocytogenes in raw milk by flow cytometric analysis of fluorescently labeled bacterial populations. The use of immunofluorescence in combination with measures of DNA content by propidium iodide labeling and size by light scattering enabled specific identification of L. monocytogenes from Streptococcus faecalis, Streptococcus agalactiae, Streptococcus uberis, Staphylococcus epidermidis, and Staphylococcus hyicus. Additional specific resolution of L. monocytogenes populations was achieved through selective enrichment of raw milk in Listeria enrichment broth. These procedures should permit the rapid screening of milk and other food samples for L. monocytogenes and eliminate many of the short-comings associated with conventional fluorescent-antibody procedures.
Fine DH, Mandel ID (1986) Indicators of periodontal disease activity : an evaluation. J Clin Periodontol 13 :533-546
It is becoming increasingly apparent that the traditional clinical criteria are inadequate for : determining active disease sites in periodontitis, monitoring quantitatively the response to therapy or measuring the degree of susceptibility to future breakdown. In an attempt to develop objective measures, a wide variety of studies have been undertaken using saliva, blood, plaque and gingival crevicular fluid (GCF) as the specimen source. Examination has included : specific bacteria and their products ; host cells and their products (enzymatic and antibacterial, both immunologic and non-immunologic) ; products of tissue injury derived from local epithelial and connective tissues and bone. Although most of the work to date has failed to provide reliable aids to the clinician, refinements in techniques for sampling and the availability of more sophisticated analytic techniques give cause for optimism. Methods proposed for detection of disease-associated bacteria in subgingival plaque vary in their sensitivity and specificity. Dark field microscopy shows some correlation with existing disease ; however, the limited specificity of this method imposes severe restrictions on its usefulness. Highly specific polyclonal and monoclonal antisera to suspected pathogens Bacteroides gingivalis and Actinobacillus actinomycetemcomitans have been developed and improved methods of identification of these microbes in plaque by ELISA immunofluorescence and flow cytometry are under development. With respect to the host response, a strong correlation between antibody patterns to specific bacteria and periodontal disease categories appears to be emerging. Although most studies have focused on serum antibody derived from peripheral blood, a shift to detection of local antibody response appears to be likely. Techniques of measurement that are exquisitely sensitive have been developed for detection of major immune recognition proteins such as antibody and complement in crevicular fluid. Research efforts attempting to correlate local antibody response to local disease activity are underway. Measurement of GCF flow rate, endotoxin, H2S, butyrate and a variety of enzymes (e.g., collagenase, arylsulfatase, B-glucuronidase) show good correlation with levels of gingivitis. In periodontitis, the most promising markers of tissue breakdown are prostaglandins of the E series, the enzymes collagenase and aspartate aminotransferase, sulfated glycosaminoglycans, osteoclastic activating factor and bone resorptive capacity of crevicular cells. Assay of the migration of crevicular leucocytes in vivo can serve as an indicator of a defect in host resistance.(ABSTRACT TRUNCATED AT 400 WORDS)
Levitt D, Zable B, Bard J (1986) Binding, ingestion, and growth of Chlamydia trachomatis (L2 serovar) analyzed by flow cytometry. Cytometry 7 :378-383
We have developed a method for quantitatively assessing binding, ingestion, and growth of Chlamydia trachomatis (L2 serovar) in several mammalian cell lines using fluorescence staining and flow cytometry. Cells were incubated with chlamydia at 4 degrees C to monitor binding ; ingestion was determined by raising the temperature to 37 degrees C for 1-4 h and removing extracellular bacteria with pronase. Growth of bacteria was measured by assessing brightly stained intracellular inclusions. Fixation with methanol prior to fluorescent staining provided the most intense specific staining with minimal background, as well as preserving cell morphology. Our data reveal relatively slow ingestion of L2 by McCoy fibroblasts (maximum ingestion by 4 h) and a sizeable population of McCoy cells (30-40% of total cells) that ingest L2 but do not permit its growth under certain infectious conditions. It was possible to correlate specific histogram patterns on the flow cytometer with fluorescent microscope observations. This system provides a means of analyzing quantitative interactions between chlamydia and individual host cells.
Mackenzie NM, Pinder AC (1986) The application of flow microfluorimetry to biomedical research and diagnosis : a review. Dev Biol Stand 64 :181-193
The application of flow cytometric or microfluorimetric techniques to the biomedical sciences is a major growth area. Flow microfluorimetry is the science of analyzing and separating populations of cells (and their sub-cellular components) using fluorescent markers (usually fluorescent antibodies or fluorescent DNA-binding dyes). Each cell is analysed individually allowing assays to be performed on small samples (less than 10,000 cells). Interesting cells can be sorted out to a high degree of purity (up to 99%). Flow microfluorimetric analysis enjoys multidisciplinary use, ranging from cell biology, microbiology and immunology to cell cycle analysis and "flow karyotyping" of cells. Examples taken from both clinical and research settings are discussed.
Massa PT, Dorries R, ter Meulen V (1986) Viral particles induce Ia antigen expression on astrocytes. Nature 320 :543-546
Recent studies have shown that gamma-interferon (IFN-gamma) induces the expression of Ia antigen on astrocytes. This observation is of immunological significance because such activated astrocytes can act as antigen-presenting cells, as demonstrated with myelin basic protein for antigen-specific encephalitogenic T-cell lines. However, the lack of lymphatic drainage in brain and the presence of the so-called blood-brain barrier restricting traffic of cells and macromolecules suggests that IFN-gamma may not be readily available, at least during the initial phases of viral infections. The question therefore arises as to whether astrocytes can be induced to express Ia antigens by other signals directly related to viral infection and possibly independent of IFN-gamma. In the present report we demonstrate that a neurotropic murine hepatitis virus induces expression of Ia antigen on astrocytes in tissue culture without infection, rendering these brain cells competent to participate directly in the immune response to a viral infection.
Suzuki T, Rogers AL, Magee PT (1986) Inter- and intra-species crosses between Candida albicans and Candida guilliermondii. Yeast 2 :53-58
Hybridization was shown to occur both between strains of the imperfect diploid yeast Candida albicans and between C. albicans and the distantly related haploid yeast Candida (Pichia) guilliermondii. Prototrophic hybrids were selected from crosses of multiply marked auxotrophic mutants of the two species. In most cases, mild ultraviolet irradiation of the C. albicans partner was required. Examination of auxotrophic markers in segregants from the hybrids indicated that recombination, rather than heterokaryon formation, had occurred in these crosses. The DNA content of the hybrids varied from diploid or aneuploid (for crosses between C. albicans and C. guilliermondii) to triploid (for C. albicans x C. albicans). It seems possible that genetic exchange analogous to this may occur in nature.